The overall goal of this project is to understand how papillomaviruses replicate their genomes. Papillomaviruses have in recent years emerged as very important causal agents in human disease. These viruses, as a part of their normal life cycle, infect and transform cells in the epithelium causing benign tumors that with a low, but significant, frequency can become malignant. A deeper understanding of the viral life cycle in general, and DNA replication in particular, is of critical importance for the understanding of the disease, its transmission, and ultimately for the development of effective therapeutic measures. We are studying the replication properties of papillomaviruses using both in vivo and cell free replication systems. We are combining thee studies with genetic and biochemical analyses of the viral proteins and sequence elements that are required for viral DNA replication. In this project we will continue our general characterization of the papillomavirus replicon with emphasis on the specific functions of the viral E1 and E2 proteins in viral DNA replication. We propose (I) to study the structure and function of the E2 transcription factor which is required for viral DNA replication; (ii) to study the structure and function of the E1 protein which is the viral initiator protein, and also how these two proteins interact with each other; (iii) to determine how, as a result of this interaction, a replication competent initiator complex is formed; and (iv) to study the involvement of the E1 and E2 proteins in stable plasmid maintenance. These studies will further our general understanding of how papillomavirus replication is initiated and controlled. In addition, the interaction between the E1 and E2 proteins presents an interesting potential target for drug intervention and these studies will significantly advance our knowledge about this interaction.